An isoleucine to valine substitution in Escherichia coli acyl carrier protein results in a functional protein of decreased molecular radius at elevated pH.

Escherichia coli acyl carrier protein (ACP) has been reported to exist in at least two distinct conformers in solution. A novel form of ACP having an increased electrophoretic mobility on polyacrylamide gel electrophoresis was noted previously during work on beta-ketoacyl-acyl carrier protein synthase II (fabF) mutants of E. coli (Jackowski, S., and Rock, C. O.(1987) J. Bacteriol. 169, 1469-1473). These workers reported that the increased electrophoretic mobility of the ACP from fabF strains occurred irrespective of prosthetic group attachment or the state of acylation of the prosthetic group. Since these workers were unable to detect a difference between the amino acid sequence of the ACP from the fabF mutants and that of wild type ACP, they suggested that the increased electrophoretic mobility was due to an unknown post-translational modification of the polypeptide chain. We have reinvestigated these mutants and report that the increased electrophoretic mobility is due to a mutation within the gene (acpP) that encodes ACP. This mutation results in substitution of isoleucine for valine 43 of ACP. Site-directed mutagenesis of a synthetic ACP gene demonstrated that the amino acid substitution at residue 43 is the cause of the increased electrophoretic mobility. Gel filtration experiments indicated that the increased electrophoretic mobility results from the more compact structure of V43I ACP at high pH. The altered residue lies within the ACP region of greatest conformational lability, and thus the V43I substitution may shift the equilibrium toward the more compact conformation(s). The disulfide-linked dimer of V43I ACP was readily formed and had an electrophoretic migration greater than the dimer of wild type ACP, suggesting that formation of ACP.ACP dimers does not require structural deformation of the protein.